A number of clinically significant, inherited disorders of hemoglobin synthesis (hemoglobinopathies) affect the structure and/or rate of synthesis of globin chains of hemoglobin. The precise molecular basis of many of these disorders is not known. Such knowledge and better knowledge of the factors controlling the expression of the normal hemoglobin genes, constitute prerequisites for the further understanding and possible future control of hemoglobinopathies such as thalassemia and sickle cell anemia. The goal of the research outlined in this proposal is to isolate normal and abnormal human globin DNA and study its fine structure and function. Human globin messenger RNA (mRNA) which is a molecular copy of the globin chain genes, can easily be obtained from human peripheral blood reticulocytes. In addition, it is possible to synthesize a radioactive DNA copy (cDNA) of globin mRNA by incubating it, in the proper conditions, with the RNA-dependent DNA polymerase of RNA tumor viruses. The cDNA can then be used as a preparative or analytical hybridization probe. Equipped with pure human globin mRNA, globin cDNA, and human cellular DNA, the following studies are contemplated: 1) Isolation of human globin DNA by various techniques utilizing fractions of human DNA enriched for globin sequences by hybridization procedures, and bacterial plasmids, which can serve as cloning and amplification vehicles. 2) Nucleotide sequence analysis of globin DNA (and intergene DNA sequences) by fingerprint analysis of P32-labeled cDNA and of isolated natural globin DNA labeled in vitro. 3) Study of the molecular mechanisms of disordered hemoglobin synthesis in nucleated thalassemic erythroid (marrow) cells, by structural analysis of thalassemic globin DNA, search for deletion in DNA of patients with beta-thalassemia, and studies of mRNA stability and mRNA processing in beta plus thalassemia.